Abstract The reported clinical activity of lurbinectedin in Small Cell Lung Cancer (SCLC) prompted us to investigate the selective mechanism of this drug in SCLC cell lines. Since lurbinectedin binds the DNA minor groove modifying DNA structure, we hypothesized that it can interfere with transcription regulation. Using SHP-77 SCLC cell line, we investigated the effect of lurbinectedin on transcriptomic profile by microarray approach, at drug concentration that reduced the cell growth rate by 50%. We found only 89 differently expressed genes (DEG), 31 up- regulated and 58 down-regulated, after 6 hours of treatment whereas there was a dramatic change in gene expression profiles after 24 hours (4529 DEGs of which 1982 up-regulated and 2547 down-regulated) and 48 hours (5457 DEGs of which 2368 up-regulated and 3089 down-regulated) of exposure. Among the genes that were clearly down-regulated at both 24 and 48 hours, there was ASCL1, a gene that encodes a transcription factor required for proper development of neuronal and pulmonary neuroendocrine cells. ASCL1 regulates stemness and cell cycle progression and it is increased in high grade neuroendocrine tumours including SCLC. Western blot analysis confirmed strong reduction of the ASCL1 protein level in SHP-77 cell line. From literature data we retrieved the ChIP-Seq target genes of ASCL1 derived from different SCLC cell lines and compared them to the list of genes at basal condition in SHP-77 cell line. Then we computed Pearson correlation between the expression of ASCL1 and these targets and filtered only strong positive correlations. Through this approach, we finally selected 213 target genes of ASCL1 many of which were coherently inhibited after 24 and 48 hours of treatment. These genes were mainly involved in MAPK and Wnt signalling pathways, both crucial for SCLC. The mechanism of transcription modulation driven by ASCL1 after lurbinectedin treatment is under investigation by ChIP-Seq technology. Moreover, work is in progress to extend our findings on others SCLC cell lines and in sub-lines in which ASCL1 was silenced. We tested whether lurbinectedin shared a similar effect on transcriptomic profile as other known drugs: by dividing up and down regulated genes and testing their correlation to the drug-related gene profiles provided by the tool Enrichr, we found that lurbinectedin has a high correlation to the topoisomerase I inhibitor class. This finding deserves further experimental verifications also in view of potential clinical interest in the combination of lurbinectedin and topoisomerase I inhibitors. Citation Format: Laura Mannarino, Ilaria Craparotta, Federica Mirimao, Nicolò Panini, Monica Lupi, Giulia Protti, Roberta Frapolli, Sergio Marchini, Maurizio D'Incalci. Lurbinectedin down-regulates ASCL1 transcription factor in Small Cell Lung Cancer (SCLC) [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference on Molecular Targets and Cancer Therapeutics; 2019 Oct 26-30; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2019;18(12 Suppl):Abstract nr LB-B13. doi:10.1158/1535-7163.TARG-19-LB-B13